Measuring the impact of maternal critical care admission on short- and longer-term maternal and birth outcomes.

PURPOSE: Factors increasing the risk of maternal critical illness are rising in prevalence in maternity populations. Studies of general critical care populations highlight that severe illness is associated with longer-term physical and psychological morbidity. We aimed to compare short- and longer-term outcomes between women who required critical care admission during pregnancy/puerperium and those who did not. METHODS: This is a cohort study including all women delivering in Scottish hospitals between 01/01/2005 and 31/12/2018, using national healthcare databases. The primary exposure was intensive care unit (ICU) admission, while secondary exposures included high dependency unit admission. Outcomes included hospital readmission (1-year post-hospital discharge, 1-year mortality, psychiatric hospital admission, stillbirth, and neonatal critical care admission). Multivariable Cox and logistic regression were used to report hazard ratios (HR) and odds ratios (OR) of association between ICU admission and outcomes. RESULTS: Of 762,918 deliveries, 1449 (0.18%) women were admitted to ICU, most commonly due to post-partum hemorrhage (225, 15.5%) followed by eclampsia/pre-eclampsia (133, 9.2%). Over-half (53.8%) required mechanical ventilation. One-year hospital readmission was more frequent in women admitted to ICU compared with non-ICU populations [24.5% (n = 299) vs 8.9% (n = 68,029)]. This association persisted after confounder adjustment (HR 1.93, 95% confidence interval [CI] 1.33, 2.81, p < 0.001). Furthermore, maternal ICU admission was associated with increased 1-year mortality (HR 40.06, 95% CI 24.04, 66.76, p < 0.001), stillbirth (OR 12.31, 95% CI 7.95,19.08, p < 0.001) and neonatal critical care admission (OR 6.99, 95% CI 5.64,8.67, p < 0.001) after confounder adjustment. CONCLUSION: Critical care admission increases the risk of adverse short-term and long-term maternal, pregnancy and neonatal outcomes. Optimizing long-term post-partum care may benefit maternal critical illness survivors.

The Receiver of the Agrobacterium tumefaciens VirA Histidine Kinase Forms a Stable Interaction with VirG to Activate Virulence Gene Expression.

The plant pathogen Agrobacterium tumefaciens carries a virulence gene system that is required for the initiation of crown gall tumors on susceptible plants. Expression of the vir genes is activated by the VirA/VirG two component regulatory system. VirA is a histidine kinase which signals the presence of certain chemicals found at the site of a plant wound. The receiver domain located at its carboxyl terminus defines VirA as a hybrid histidine kinase. Here, we show that the VirA receiver interacts with the DNA-binding domain of VirG. This finding supports the hypothesis that the receiver acts as a recruiting factor for VirG. In addition, we show that removal of the VirA receiver allowed vir gene expression in response to glucose in a dose dependent manner, indicating that the receiver controls VirG activation and suggesting that the supplementary ChvE-sugar signal increases this activity.

Role of the VirA histidine autokinase of Agrobacterium tumefaciens in the initial steps of pathogenesis.

Histidine kinases serve as critical environmental sensing modules, and despite their designation as simple two-component modules, their functional roles are remarkably diverse. In Agrobacterium tumefaciens pathogenesis, VirA serves with VirG as the initiating sensor/transcriptional activator for inter-kingdom gene transfer and transformation of higher plants. Through responses to three separate signal inputs, low pH, sugars, and phenols, A. tumefaciens commits to pathogenesis in virtually all flowering plants. However, how these three signals are integrated to regulate the response and why these signals might be diagnostic for susceptible cells across such a broad host-range remains poorly understood. Using a homology model of the VirA linker region, we provide evidence for coordinated long-range transmission of inputs perceived both outside and inside the cell through the creation of targeted VirA truncations. Further, our evidence is consistent with signal inputs weakening associations between VirA domains to position the active site histidine for phosphate transfer. This mechanism requires long-range regulation of inter-domain stability and the transmission of input signals through a common integrating domain for VirA signal transduction.

The integrity of the periplasmic domain of the VirA sensor kinase is critical for optimal coordination of the virulence signal response in Agrobacterium tumefaciens.

The plant pathogen Agrobacterium tumefaciens responds to three main signals at the plant-bacterium interface: phenolics, such as acetosyringone (AS), monosaccharides, and acidic pH (∼5.5). These signals are transduced via the chromosomally encoded sugar binding protein ChvE and the Ti plasmid-encoded VirA/VirG two-component regulatory system, resulting in the transcriptional activation of the Ti plasmid virulence genes. Here, we present genetic and physical evidence that the periplasmic domain of VirA dimerizes independently of other parts of the protein, and we examine the effects of several engineered mutations in the periplasmic and transmembrane regions of VirA on vir-inducing capacity as indicated by AS sensitivity and maximal level of vir-inducing activity at saturating AS levels. The data indicate that helix-breaking mutations throughout the periplasmic domain of VirA or mutations that reposition the second transmembrane domain (TM2) of VirA relieve the periplasmic domain's repressive effects on the maximal activity of this kinase in response to phenolics, effects normally relieved only when ChvE, sugars, and low pH are also present. Such relief, however, does not sensitize VirA to low concentrations of phenolics, the other major effect of the ChvE-sugar and low pH signals. We further demonstrate that amino acid residues in a small Trg-like motif in the periplasmic domain of VirA are crucial for transmission of the ChvE-sugar signal to the cytoplasmic domain. These experiments provide evidence that small perturbations in the periplasmic domain of VirA can uncouple sugar-mediated changes in AS sensitivity from the sugar-mediated effects on maximal activity.

The receiver domain of hybrid histidine kinase VirA: an enhancing factor for vir gene expression in Agrobacterium tumefaciens.

The plant pathogen Agrobacterium tumefaciens expresses virulence (vir) genes in response to chemical signals found at the site of a plant wound. VirA, a hybrid histidine kinase, and its cognate response regulator, VirG, regulate vir gene expression. The receiver domain at the carboxyl end of VirA has been described as an inhibitory element because its removal increased vir gene expression relative to that of full-length VirA. However, experiments that characterized the receiver region as an inhibitory element were performed in the presence of constitutively expressed virG. We show here that VirA's receiver domain is an activating factor if virG is expressed from its native promoter on the Ti plasmid. When virADeltaR was expressed from a multicopy plasmid, both sugar and the phenolic inducer were essential for vir gene expression. Replacement of wild-type virA on pTi with virADeltaR precluded vir gene induction, and the cells did not accumulate VirG or induce transcription of a virG-lacZ fusion in response to acetosyringone. These phenotypes were corrected if the virG copy number was increased. In addition, we show that the VirA receiver domain can interact with the VirG DNA-binding domain.

Challenges of major obstetric haemorrhage.

Every minute of every day, a woman dies in pregnancy or childbirth. The biggest killer is obstetric haemorrhage, the successful treatment of which is a challenge for both the developed and developing worlds. The presence of an attendant at every birth and access to emergency obstetric care are key to reducing maternal morbidity and mortality in the developing world while resource-rich countries have a rising caesarean section rate with its consequential effect on the incidence of abnormal placentation and its link with peripartum hysterectomy. Management of obstetric haemorrhage involves early recognition, assessment and resuscitation. Various methods are available to try to stop the bleeding - from pharmacological methods to aid uterine contraction (e.g., oxytocinon, ergometrine and prostaglandins) to surgical methods to stem the bleeding (e.g., balloon tamponade, compression sutures or arterial ligation). Interventional radiology can be used if placenta accreta is suspected. Cell salvage has been introduced into obstetrics relatively recently in an attempt to reduce allogeneic transfusion.

Strategies to manage major obstetric haemorrhage.

PURPOSE OF REVIEW: Haemorrhage remains a cause of significant maternal morbidity and mortality. This review summarizes the prevention, management and treatment of obstetric haemorrhage and highlights recent advances and developments. RECENT FINDINGS: Postpartum haemorrhage is the most common cause of major obstetric haemorrhage and is usually due to uterine atony. Pharmacological treatment has not altered much in recent years with oxytocin and ergometrine remaining first-line options. Although controversy surrounds its advantages over other uterotonics, the use of misoprostol has been increasing, especially in resource-poor countries. Placenta accreta is becoming more common, a sequelae to the rising caesarean section rate. Interventional radiology may reduce blood loss in these cases. Uterine compression sutures, intrauterine tamponade balloons and cell salvage have all made their debut in the last decade. SUMMARY: Accurate diagnosis and appropriate management of obstetric haemorrhage can reduce maternal morbidity and mortality. This review outlines the current evidence.

The plant signal salicylic acid shuts down expression of the vir regulon and activates quormone-quenching genes in Agrobacterium.

Agrobacterium tumefaciens is capable of transferring and integrating an oncogenic T-DNA (transferred DNA) from its tumor-inducing (Ti) plasmid into dicotyledonous plants. This transfer requires that the virulence genes (vir regulon) be induced by plant signals such as acetosyringone in an acidic environment. Salicylic acid (SA) is a key signal molecule in regulating plant defense against pathogens. However, how SA influences Agrobacterium and its interactions with plants is poorly understood. Here we show that SA can directly shut down the expression of the vir regulon. SA specifically inhibited the expression of the Agrobacterium virA/G two-component regulatory system that tightly controls the expression of the vir regulon including the repABC operon on the Ti plasmid. We provide evidence suggesting that SA attenuates the function of the VirA kinase domain. Independent of its effect on the vir regulon, SA up-regulated the attKLM operon, which functions in degrading the bacterial quormone N-acylhomoserine lactone. Plants defective in SA accumulation were more susceptible to Agrobacterium infection, whereas plants overproducing SA were relatively recalcitrant to tumor formation. Our results illustrate that SA, besides its well known function in regulating plant defense, can also interfere directly with several aspects of the Agrobacterium infection process.

Culture and maintenance of Agrobacterium strains.

As aerobic chemoorganotrophs, most Agrobacterium strains will grow on a wide range of complex and defined media. Methods commonly used for the culture and storage of other chemoorganotrophs will usually work for agrobacteria as well. Problems with culture or strain maintenance will occur more frequently because of careless technique than because of strain difficulties. Here we describe a few of the complex and defined media that have been successfully used in the growth of agrobacteria including some that are semiselective for agrobacteria. Finally, we present methods suitable for short- and long-term storage of Agrobacterium strains.

Three methods for the introduction of foreign DNA into Agrobacterium.

The genetic manipulation of Agrobacterium tumefaciens is used to facilitate studies of bacterial gene functions or as a first step in introducing genetic material into transformable plant cells through the use of T-DNA binary vectors. Three methods are commonly used. Transformation with purified plasmid can be done with either electroporation or a simple freeze/thaw transformation method. Alternatively, a mobilizable plasmid can be placed into Agrobacterium using the triparental mating method. Here we present three detailed protocols for Agrobacterium strain construction using electroporation, the freeze/thaw method of transformation, and triparental mating.

Nucleic acid extraction from Agrobacterium strains.

Agrobacterium is routinely used as a tool for moving genetic constructs into plant cells. The successful use of Agrobacterium as a tool for the genetic engineering of plant cells often requires the manipulation and analysis of nucleic acids present in recombinant Agrobacterium strains. Here we present dependable methods for the isolation of genomic (total) DNA, mega-plasmid DNA, shuttle or binary plasmid DNA, and RNA. In addition, we provide a simple method for the electronic transfer of shuttle plasmids from Agrobacterium to E. coli for use when their low copy number in Agrobacterium impedes plasmid isolation from that strain.

Intersubunit complementation of sugar signal transduction in VirA heterodimers and posttranslational regulation of VirA activity in Agrobacterium tumefaciens.

The VirA/VirG two-component regulatory system of Agrobacterium tumefaciens regulates expression of the virulence (vir) genes that control the infection process leading to crown gall tumor disease on susceptible plants. VirA, a membrane-bound homodimer, initiates vir gene induction by communicating the presence of molecular signals found at the site of a plant wound through phosphorylation of VirG. Inducing signals include phenols, monosaccharides, and acidic pH. While sugars are not essential for gene induction, their presence greatly increases vir gene expression when levels of the essential phenolic signal are low. Reception of the sugar signal depends on a direct interaction between ChvE, a sugar-binding protein, and VirA. Here we show that the sugar signal received in the periplasmic region of one subunit within a VirA heterodimer can enhance the kinase function of the second subunit. However, sugar enhancement of vir gene expression was vector dependent. virA alleles expressed from pSa-derived vectors inhibited signal transduction by endogenous VirA. Inhibition was conditional, depending on the induction medium and the virA allele tested. Moreover, constitutive expression of virG overcame the inhibitory effect of some but not all virA alleles, suggesting that there may be more than one inhibitory mechanism.